Asthma is associated with variable responses to corticosteroids. There have been no functional studies analyzing airway microbiome effects on corticosteroid responsiveness in asthmatics. National and international guidelines (NIH publication no 07-4051. Bethesda, Md.: National Institutes of Health National Heart Lung and Blood Institute; 2007; Global strategy for asthma management and prevention 2009 (update): Global Initiative for Asthma; 2009) recommend the use of corticosteroids to control airway inflammation in persistent asthma Clinical studies, however, demonstrate highly variable responses to corticosteroid therapy with up to 45% of patients not having a clinical or physiologic response to inhaled corticosteroids (ICS) (Malmstrom K et al., Ann Intern Med 1999; 130:487-95; Martin R J et al., J Allergy Clin Immunol 2007; 119:73-80; Barnes P J et al., Lancet 2009; 373:1905-17), and up to 25% of patients not responding to oral corticosteroids (Leung D Y et al., J Allergy Clin Immunol 2003; 111:3-22; quiz 3). These corticosteroid resistant (CR) patients have increased airway inflammation (Leung D Y et al., J Allergy Clin Immunol 2003; 111:3-22; quiz 3; Leung D Y et al., J Exp Med 1995; 181:33-40) despite treatment with oral steroids, and airway remodeling (Goleva E et al., J Allergy Clin Immunol 2007; 120:1065-72) contributing to the severity of asthma. Given variable responses to corticosteroid therapy in asthmatics, alternative therapeutics targeting disease causality are needed for personalized treatment of asthma (Drazen J M., J Allergy Clin Immunol 2012; 129:1200-1).
Glucocorticoids (GCs) are the most potent anti-inflammatory drugs used for treatment of asthma and other chronic inflammatory or autoimmune diseases. Up to 20% of asthmatics remain refractory to CS or glucocorticoid (GC) therapy—these patients are referred to as steroid resistant (SR) or steroid insensitive asthmatics (McManus R., J Endocrinol 2003; 178:1-4). SR asthmatics are characterized by increased airway inflammation that cannot be inhibited by CS treatment. The role of race, smoking, obesity, vitamin D level, allergens, and infection in steroid resistance is under active investigation (Leung D Y et al., J Allergy Clin Immunol 2003; 111:3-22; quiz 23; Althuis M D et al., J Asthma 1999; 36:257-264; Sutherland E R et al., Am J Respir Crit Care Med 2010; 181:699-704). In addition, endotoxin exposure has recently been identified as an important factor that alters cellular response to CS (Yang M et al., J Immunol 2009; 182:5107-5115; Goleva E et al., J Allergy Clin Immunol 2008; 122:550-559 e553; Bhaysar P et al., Thorax 2008; 63:784-790).
Endotoxin or lipopolysaccharide (LPS), a component of the outer membrane of Gram-negative bacteria, has been implicated in innate immune responses by inducing activation and release of proinflammatory cytokines, nitric oxide, reactive oxygen species and other cell mediators from monocytes and macrophages (Rossol M et al., Crit Rev Immunol 2011; 31:379-446). Exposure to endotoxin has been associated with asthma exacerbation (Liu A H., Paediatr Respir Rev 2004; 5 Suppl A:S65-71). Several studies have demonstrated that higher exposure to house dust endotoxin is related to asthma severity (Michel 0 et al., Am J Respir Crit Care Med 1996; 154:1641-1646; Rizzo M C et al., Pediatr Allergy Immunol 1997; 8:121-126P). Previous studies also indicate that exposure of monocytes to LPS induces cellular steroid resistance (Goleva E et al., J Allergy Clin Immunol 2008; 122:550-559 e553).
Mammalian cells respond to LPS stimulation through a series of protein interactions, including engagement of LPS binding protein (LBP), CD14, and Toll-like receptor (TLR) 4 (Gioannini T L, et al. J Biol Chem 2007; 282:7877-7884), its cell membrane receptor. Intracellular signaling mediated by LPS/TLR4 involves binding of a series of adaptor molecules, including TRIF, TRAM, TIRAP, BTK and MyD88, and leads to sequential kinase phosphorylation (Fitzgerald K A et al., J Exp Med 2003; 198:1043-1055). Mitogen activated protein kinases (MAPK)s are the final kinases in this cascade and include p38 MAPK (p38), p42/44 MAPK (ERK) and c-Jun NH2-terminal kinase (JNK) (Guha M, Mackman. Cell Signal 2001; 13:85-94). Stimulation of human monocytes with LPS has been shown to result in the phosphorylation and activation of p38, ERK and JNK (Carter A B et al., Am J Respir Cell Mol Biol 1999; 20:751-758; Lim W et al., J Immunol 2005; 175:5690-5700). Several studies have demonstrated that MAPK pathways are involved in activation of transcription factors, such as NF-κB and AP-1 (Rawadi G et al., J Immunol 1999; 162:2193-2203; Oeckinghaus A et al., Nat Immunol 2011; 12:695-708); these transcription factors play a critical role in LPS-induced expression of proinflammatory genes, such as TNF-α, IL-10, IL-6, IL-8, MCP-1, E-selectin, VCAM-1 and ICAM-1.
Cytoplasmic glucocorticoid receptor (GCR) mediates cellular response to GCs. Activated GCR translocates to the cell nuclei and acts as a transcriptional factor. GCR can inhibit pro-inflammatory MAPK signaling by inducing nuclear mitogen activated kinase phosphatase (MKP1) expression (Liu Y et al., Nat Rev Immunol 2007; 7:202-212; Lasa M et al., Mol Cell Biol 2002; 22:7802-7811). At the same time, GCR activity is subject to kinase modulation, activated MAPKs can inhibit GCR function via phosphorylation that will inhibit GCR nuclear translocation in response to GC treatment, cause the GCR to return to the cytoplasm or modify GCR transcriptional activity (Ismaili N et al., Ann N Y Acad Sci 2004; 1024:86-101; Galliher-Beckley A J et al., IUBMB Life 2009; 61:979-986).
In addition, 16s rRNA gene sequence analysis of the airway microbiota has demonstrated diversified microbial communities in the airways of asthmatics (Hilty M et al., PLoS One 2010; 5:e8578; Huang Y J et al., J Allergy Clin Immunol 2011; 127:372-81 e1-3). Patients with asthma showed significantly higher bacterial diversity due to expansion of pathogenic bacteria and greater bacterial burden compared with healthy controls. Moreover, airway microbiome composition and greater bacterial diversity significantly correlates with bronchial hyperresponsiveness, including the relative abundance of bacterial families within Proteobacteria. The functional impact of airway microbiome and its contribution to pathophysiologic processes in asthma were not previously known.